[unreadable] The amino acid sequence of a protein determines its fold. However, small changes such as single amino acid substitutions or deletions can cause misfolding, and subsequent aggregation. Protein misfolding directly causes several devastating human diseases. Molecular chaperones assist the folding of many proteins, especially those prone to misfolding and aggregation. However, how chaperones distinguish between incorrectly and productively folding proteins is unclear. The mechanism of substrate specificity of the major bacterial chaperone complex, GroEL/ES, will be investigated. The major aim of this proposal is to select and characterize mutants of GroEL/ES that more efficiently fold substrate proteins. The coat protein of phage P22 will be the model substrate used to isolate the chaperone mutants because several single amino acid substitutions convert coat protein to become a substrate for GroEL/ES, though with limited efficiency. GroEL/ES mutants will be selected that can fold the coat protein mutants at high temperatures. The ability of the GroEL/ES mutants to fold a variety of substrate proteins will be determined both in vivo and in vitro to gain a generalized understanding of how these chaperone proteins recognize substrate polypeptides. [unreadable] [unreadable] [unreadable]